1. Field of the Invention
The present invention relates to a novel analogue antibiotic peptide derived from a CM-MA peptide and a use thereof.
2. Description of the Related Art
Bacterial infection is the most common and general but sometimes an incurable cause of disease. Unfortunately, bacteria gain resistance against antibiotics owing to the over-use of antibiotics. In fact, gaining resistance of bacteria against a new antibiotic agent outruns the development of a new antibiotic analogue. For example, lethal bacteria such as Enterococcus faecalis, Mycobacterium tuberculosis, and Pseudomonas aeruginosa have grown their resistance against almost all antibiotics known so far (Stuart B. Levy, Scientific American, 1998, 46-53).
Tolerance against antibiotics is different from resistance against antibiotics. Tolerance was first identified in Pneumococcus sp. in 1970s and provided an important clue for the mechanism of penicillin (Tomasz et al., Nature, 1970, 227, 138-140). Bacteria that show tolerance stop their growth in the presence of antibiotics but are not dead. Tolerance is generated when the autolytic enzymes such as autolysin are not activated because of the antibiotics inhibiting the enzyme involved in the synthesis of cell wall. This phenomenon explains that penicillin activates endogenous hydrolytic enzyme so as to kill bacteria and reversely bacteria inhibit the activity of the enzyme so as to survive in the presence of antibiotics.
It is a clinically very important fact that bacteria have tolerance against various antibiotics. If it is not possible to kill resistant bacteria, the treatment effect of an antibiotic agent for clinical infection would be diminished (Handwerger and Tomasz, Rev. Infec. Dis., 1985, 7, 368-386). Being tolerant is the first step of being resistant. So, after all the treatment with antibiotics, there are still strains survived. Such strains acquire a new genetic element that shows resistance against antibiotics so that they keep growing in the presence of the antibiotics. Actually all the strains showing resistance are confirmed to have tolerance as well (Liu and Tomasz, J. Infect. Dis., 1985, 152, 365-372). Therefore, it is required to develop a novel antibiotic agent that can kill the strain displaying resistance against antibiotics.
Tolerance is acquired by two different pathways at large in the aspect of mechanism. Phenotypic tolerance is the one that is generated in all of bacteria when the growth rate decreases (Tuomanen E., Revs. Infect. Dis., 1986, 3, S279-S291) and genetic tolerance is the other that is generated by genetic mutation and identified only in some specific bacteria. The basic phenomenon of the two types of tolerance is down regulation of autolysin activity. The down regulation of autolysin activity caused by phenotypic tolerance is temporary but the down regulation of autolysin activity caused by genetic tolerance is permanent because of the mutation that changes the pathway to regulate cell lysis. The simplest genetic tolerance is the defect in autolysin. The deficiency of autolysin did not produce such a strain that had tolerance by some unknown reasons and rather clinical tolerance was observed in the regulation of autolysin (Tuomanen et al., J. infect. Dis., 1988, 158, 36-43).
As explained hereinbefore, it is necessary to develop a novel antibiotic agent in order to cope such bacteria that show resistance against antibiotics. It is more important to develop a novel antibiotic agent that works independently from the autolysin activity. It is thus required to provide a novel antibiotic agent for the treatment of bacterial infection and inflammation.
Bacteria can kill neighbor bacteria by synthesizing peptides or small organic molecules, which are called bacteriocin. The bacteriocin is classified into three groups according to the structure; lantibiotics, nonlantibiotics, and those secreted by signal peptide (Cintas et al., J. Bad., 1998, 180, 1988-1994). Insects and animals can produce endogenous peptide antibiotics (Bevins et al., Ann. Rev. Biochem., 1990, 59, 395-414), which are also divided into three groups according to the structure. The first group is cysteine-rich β-sheet peptides, the second group is α-helical amphiphilic molecules, and the third group is proline-rich peptides (Mayasaki et al., Int. J. Antimicrob. Agents, 1998, 9, 269-280). These antibiotic peptides play an important role in host defense and innate immune system (Boman, H. G., Cell, 1991, 65, 205; Boman, H. G., Annu. Rev. Microbiol., 1995, 13, 61). These anti-bacterial peptides display different structures generated by different amino acid sequences. The most frequently observed antibacterial peptide has the cysteine free amphiphilic α-helical structure like cecropin, identified mostly in insects.
The antibacterial activity of the amphiphilic peptide has been most studied among those peptides, based on which attempts have been made to develop an antibacterial agent. The amphiphilic peptides reported so far are magainin 2 (MA), cecropin A (CA), and melittin (ME), etc.
The cecropin family amphiphilic peptide was first identified in drosophila. Later, it was also found in silkworm pupa and hog small intestine. Cecropin A exhibits high antibacterial activity but weak anti-fungal activity and anticancer activity (Boman, H. G. and Hultmark, D., Annu. Rev. Microbiol., 1987, 41, 103). Magainin 2 peptide has no cytotoxicity and displays anti-fungal, anticancer, and anti-protozoan activities along with anti-bacterial activity (Zasloff, M., Proc. Natl. Acad. Sci. USA, 1987, 84, 5449). It is also known that a synthetic peptide having an excellent anti-bacterial, anti-fungal, or anti-cancer activity can be prepared as a conjugation peptide produced by conjugating some parts of sequences of the two peptides above (Chan, H. C., et al., FEBS Lett., 1989, 259, 103; Wade, D., et al., Int. J. Pept. Prot. Res., 1992, 40, 429).
The present inventors tried to produce a novel synthetic peptide with the improved antibacterial activity from the conventional peptides reported to have an antibacterial activity. As a result, the inventors succeeded in synthesizing the novel antibiotic peptides represented by SEQ. ID. NO: 2˜NO: 7 (CMA1˜CMA 6) by using a CA-MA antibiotic peptide, in which amphiphilic cecropin A (CA) and magainin 2 (MA) were conjugated, as a template. The synthesized antibiotic peptides displayed antibacterial activity against gram-positive and gram negative bacteria and at the same time displayed low cytotoxicity to human red blood cells and human normal cell lines (HaCaT). Therefore, the synthesized novel antibiotic peptides were confirmed to be effectively used as an active ingredient of an antibiotic agent, a cosmetic composition, a food additive, a feed additive, a biological pesticide, and a quasi-drug, leading to the completion of the present invention.